Studies of the Mechanism of Cation Transport

Abstract

SUMMARY.The distribution und activity of adenosine-triphosphatase in homogenates and subfractions of guinea pig kidney cortex has bt-en examined. A portion of the activity associated with particulate fractions could IH; furlhtr stimulated, in the presence of magnesium ions, by the combined addition of sodium and potassium ions but not by the addition of either cation alone or in combination with choline.Treatment of the heavy microsome subfraction with deoxycholate resulted in a large increase in the specific activity of this enzyme, which now re(|uired sodium and potassium ions in a ratio of 4 : 1 for over 85 p.c. of its total activity. Under such conditions of cation stimulation, optimum pH was found to be between 7-5 and 8-0, and the enzyme also exhibited considerable temperature dependence and was stable up to 55° C.The cardiac glycoside Ouabain completely inhibited that portion of enzyme activity stimulated by monovalent cittions but was without effect on the activity observed in the presence of niiiKnesium ions alone.The effect of other reagents and hormones on the cation stimulated adenosine-triphosphatase activity was also examined, with a discussion of the possible significance of this enzyme in active cation transport.INTRODUCTION.

STUDIES ON Na+‐K+‐STIMULATED ATPase OF HUMAN BRAIN*

“…The Na+-K+-stimulated ATPase in the salt-insoluble microsomal fraction from human grey matter is 3 times as active as a similar preparation from white matter. The pH optimum of 7.5 for the Na+-K+-ATPase activity of human brain is similar to that found for the analogous enzyme prepared from human muscle (SAMAHA and GERGELY, 1966) and other animal tissues (SKOU, 1957; DEUL and MCILWAIN, 1961 ;SCHWARTZ, LASETER and KRAINTZ, 1963;LEE and Yu, 1963;CHARNOCK and POST, 1963). In addition, the inhibition of the Na+-K+-ATPase activity by PCMB and by high concentrations of quinidine and oligomycin is similar to that shown by analogous enzymes from other tissues (CHARNOCK and POST, 1963;LASETER, 1963, GLYNN, 1963; VAN GRONINGEN and SLATER, 1963;WHITTAM, WHEELER and BLAKE, 1964;SAMAHA and GERGELY, 1966).…”

“…The pH optimum of 7.5 for the Na+-K+-ATPase activity of human brain is similar to that found for the analogous enzyme prepared from human muscle (SAMAHA and GERGELY, 1966) and other animal tissues (SKOU, 1957; DEUL and MCILWAIN, 1961 ;SCHWARTZ, LASETER and KRAINTZ, 1963;LEE and Yu, 1963;CHARNOCK and POST, 1963). In addition, the inhibition of the Na+-K+-ATPase activity by PCMB and by high concentrations of quinidine and oligomycin is similar to that shown by analogous enzymes from other tissues (CHARNOCK and POST, 1963;LASETER, 1963, GLYNN, 1963; VAN GRONINGEN and SLATER, 1963;WHITTAM, WHEELER and BLAKE, 1964;SAMAHA and GERGELY, 1966). In other studies, a consistent observation has been made concerning the substitution of other monovalent cations, namely that K+ is a nonspecific requirement since it can be replaced by NH,+, Rb+, Cs+ and Li+ (in descending order of effectiveness) but that Na+ is specific since it cannot be replaced by any other cations (SKOU, 1962; LANDON and NORRIS, 1963;RENDI and UHR, 1964;SAMAHA and GERGELY, 1966).…”

Salt-Stimulated Adenosine Triphosphatases from Carrot, Beet, and Chara Australis

“…For routine m.easurement of salt-stimulated ATPase, assays were carried out in the preSence and absence of 0 ·IM KCl. Orthophosphate -liberated by the ATPase was measuredcolorimetrically with acid molybdate and amidol as described by Charnock and Post (1963); rates observed were constant for 15 min with or without potassium chloride. In preliminary studies the colorimetric assay was confirmed by chromatography of the products in isobutyric acid-O;5N NH40H (2,: 1 v/v) and spectrophotometric analysis of ATP, ADP, and AMP.…”

“…Poulik 1957). After electrophoresis for 4 hr at 40 mA in an apparatus cooled to 0-4°C the gel (cross section 10 cm by 1 cm) was sliced with a fine nylon thread (for details of methods see Clare, Flentje, and Atkinson 1968). One-half was stained for ATPase by soaking in a filtered solution of 2 mM disodium ATP-3 mM Pb(N03)2-50 mM acetate (Tris, pH 4·5) for 1-2 hr, rinsed thoroughly in water, and soaked in 0·1 % ammonium sulphide.…”

Effects of Serum Proteins in vitro on the Synthesis of Rapidly Labelled Ribonucleic Acid and on the Polysome Pattern of Yoshida Ascites-Hepatoma Cells

“…To examine this further, hydroxylamine (036M) was substituted for K+ (20mM) in three assays of ATPase activity with a standard chemical assay system for the liberation of inorganic phosphate from non-radioactive ATP (2mM) in the presence of 80mm-Na+ (Charnock & Post, 1963b (values are the means of three experiments, assays in duplicate). In every experiment addition of 01 mm-ouabain decreased the liberation of phosphate to that found with Na+ alone.…”